DESCRIPTION: Basic and clinical research suggests there are extensive bidirectional interactions between circadian rhythms and addiction. Disruptions to the circadian system, either by environmental or genetic perturbation, may increase the vulnerability to addiction, while chronic drug use leads to circadian disruptions that persist during abstinence and may contribute to relapse. Although these relationships are intriguing, very little is known about the molecular mechanisms underlying the relationship between the circadian system and the transition to addiction. Animal studies have demonstrated that chronic exposure to cocaine leads to alterations in the expression and function of specific circadian genes (i.e., components of the molecular clock) in the mesolimbic dopamine reward system. A major region of convergence for reward circuitry and a key substrate that regulates drug reward and motivation is the nucleus accumbens (NAc). The NAc is comprised of mostly two specific subtypes of medium spiny neurons (MSNs) that predominantly express either dopamine 1 or 2 receptors (D1+ or D2+). These two subtypes of MSNs have distinct roles in the regulation of cocaine reward behaviors, although the molecular mechanisms underlying these differences remain unclear. Recent human genetics studies have identified variants in the gene coding for the circadian transcription factor neuronal PAS domain protein 2 (NPAS2) associated with psychiatric disorders that are highly comorbid with addiction disorders. We have identified a novel role of NPAS2 in the regulation of cocaine reward via activity in D1+ MSNs of the NAc. The R21 aims are as follows: 1) Leverage CRISPR/Cas9 technology to generate split Cre mice that enable us to use intersectional genetics approaches to target D1+ or D2+ MSNs specifically in the striatumR and 2) Generate NPAS2Qdeficient mice exclusively in striatal D1+ or D2+ MSNs to investigate the cell type specific molecular mechanisms regulating reward and motivation (R33 phase). The R33 aims will characterize these mice by 1) investigating the role of Npas2 in the regulation of circadian regulation of cocaine conditioned reward (conditioned place preference) and self-administration 2) NPAS2Q mediated circadian transcription on cocaine-induced dendritic plasticity in the NAcR and 3) elucidating the cell type specific molecular mechanisms of NPAS2 regulation of cocaine reward using confocal microscopy, FACs, RNA seq, and integrative analyses with preliminary ChIPseq data. These studies will further clarify the role of the molecular clock in the transition to addiction, and importantly, provide the broader scientific community with novel transgenic mice to further investigate the molecular mechanisms of 'direct' and 'indirect' pathway regulation of drug reward and addiction phenotypes. These studies will leverage CRISPR/Cas9 technologies and advanced molecular and behavioral approaches to study a novel mechanism of circadian regulation of addiction behaviors.